International Patent Classification: A 61 K 31/70, C 07 H 17/08
The present invention relates to new tylosin derivatives, new synthetic products of the macrolide class exhibiting antimicrobial activity. It particularly relates to 3-deoxy-3-oxo-desmycosin derivatives of the formula (I) 
wherein
R represents CHO or CH(OCH3)2, R1 and R2 represent H or acetyl, R3 represents H or OH, R4 represents N(CH3)2 or Nxe2x80x94O(CH3)2, the line - - - represents a single or a double bond, the line . . . represents  or a double or a single bond, and the line  represents a double or a single bond, and to derivatives of 3-deoxy-2,3-didehydro-desmycosin of the formula II 
wherein
R represents CHO or CH(OCH3)2, R1 represents H or OH, R2 represents N(CH3)2 or Nxe2x80x94O(CH3)2, the line - - - represents a double or a single bond, the line . . . represents  or a single bond, and the  line represents a double or a single bond, and to a process for the preparation thereof.
It is known that 13-hydroxy derivatives of tylosin have been prepared by a reductive opening of the oxirane ring (A. Narandja, SI 9700281). It is also known that 10,11,12,13-tetrahydro derivatives of tylosin have been prepared by catalytic hydrogenation of tylosin (A. Narandja, EP 287082 B3). It is also known that 3-deoxy-2,3-didehydro derivatives of tylosin (S. Kageyama, Bull. Chem. Jpn. 65, 3405,1992) as well as 3-deoxy-3-oxo derivatives of 6-O-methyl-erythromycin (A. Agouridas, J. Med. Chem. 41, 4080, 1998) have been prepared.
According to the known prior art, however, neither 3-deoxy-3oxo-derivatives of tylosin class nor 3-deoxy-2,3-didehydro derivatives with exchanged left side of the molecule in C-10 to C-13 positions and processes for the preparation thereof have been described so far.
It has been found that derivatives of 3-deoxy-3-oxo-desmycosin of the formula I 
wherein
R represents CHO or CH(OCH3)2, R1 and R2 represent H or acetyl, R3 represents H or OH, R4 represents N(CH3)2 or Nxe2x80x94O(CH3)2, the line - - - represents a single or a double bond, the line . . . represents  or a double or a single bond, and the line  represents a double or a single bond, and 3-deoxy-2,3-didehydro-derivatives of the formula II 
wherein
R represents CHO or CH(OCH3)2, R1 represents H or OH, R2 represents N(CH3)2 or Nxe2x80x94O(CH3)2, the line - - - represents a double or a single bond, the line . . . represents  or a single bond, and the line  represents a double or a single bond, can be prepared starting from a compound of the formula III 
wherein R represents H or SO2CH3 and the line - - - represents a double or a single bond.
According to the present invention the compound of formula III, wherein R represents H and the line - - - represents a double bond, is subjected
A/ to an oxidation reaction in a solution of methylene chloride in the presence of 15-28 equivalents of DMSO, 8-14 equivalents of N-(3-dimethylaminopropyl)-Nxe2x80x2-ethyl carbodiimide hydrochloride and 8-14 equivalents of pyridine trifluoroacetate within 2-6 hours at a temperature of 10-25xc2x0 C., whereupon the obtained compound of the formula I, wherein R represents CH(OCH3)2, R1 and R2 represent acetyl, R3 represents H, R4 represents N(CH3)2, the lines - - - and . . . represent double bonds and the line  represents a single bond,
is optionally subjected
B/ to a methanolysis at reflux temperature within 4-6 hours and the obtained compound of the formula I, wherein R represents CH(OCH3)2, R1 and R3 represent H, R2 represents acetyl, R4 represents N(CH3)2, the lines - - - and . . . represent double bonds and the line  represents a single bond,
is optionally subjected
B1/ to an alkaline methanolysis in a mixture of methanol and 25% NH4OH (2:1) at 5xc2x0 C. for a period of 48-60 hours, whereupon the obtained compound of the formula I wherein R represents CH(OCH3)2, R1, R2 and R3 represent H, R4 represents N(CH3)2, the lines - - - and . . . represent double bonds and the line  represents a single bond,
is optionally subjected
B2/ to a hydrolysis in a mixture of acetonitrile and 1% trifluoroacetic acid (2:3) within 2 hours at room temperature, giving a compound of the formula I, wherein R represents CHO, R1, R2 and R3 represent H, R4 represents N(CH3)2, the lines - - - and . . . represent double bonds and the line  represents a single bond,
or optionally,
a compound of the formula I, wherein R represents CH(OCH3)2, R1 and R2 represent acetyl, R3 represents H and R4 represents N(CH3)2, the lines - - - and . . . represent double bonds and the line  represents a single bond,
is subjected
C/ to a catalytic hydrogenation reaction in the presence of 2-5% Pd/C (w/w) at room temperature within 5-8 hours at a hydrogen pressure of 0.3-0.5 MPa, whereupon the obtained compound of the formula I, wherein R represents CH(OCH3)2, R1 and R2 represent acetyl, R3 represents H, R4 represents N(CH3)2, and the lines - - - , . . . and  represent single bonds, can be optionally subjected to methanolysis or alkaline methanolysis reactions in the manner described under B or B1;
or optionally, it is subjected
D/ to an oxidation reaction in a methylene chloride solution in the presence of 3-6 equivalents of m-chloroperbenzoic acid within 6-10 hours at room temperature, giving a compound of the formula I, wherein R represents CH(OCH3)2, R1 and R2 represent acetyl, R3 represents H, R4 represents Nxe2x80x94O(CH3)2, the line - - - represents a double bond, the line . . . represents  and the line  represents a single bond,
or optionally,
a compound of the formula I, wherein R represents CH(OCH3)2, R1 and R3 represent H, R2 represents acetyl, R4 represents Nxe2x80x94(CH3)2, the lines - - - and . . . represent double bonds and the line  represents a single bond, is subjected to the oxidation reaction in the manner described under D, and the obtained compound of the formula I, wherein R represents CH(OCH3)2, R1 and R3 represent H, R2 represents acetyl, R4 represents Nxe2x80x94O(CH3)2, the line - - - represents a double bond, the line . . . represents  and the line  represents a single bond,
is optionally subjected
to the catalytic hydrogenation reaction in the manner described under C, giving a compound of the formula I, wherein R represents CH(OCH3)2, R1 and R3 represent H, R2 represents acetyl, R4 represents N(CH3)2, the line - - - represents a single bond, the line . . . represents  and the line  represents a single bond,
is optionally subjected
E/ to a reduction reaction with Zn-powder in a solution of EtOH and a 10% aqueous NH4OH solution (1:2) under maintaining the pH-value of 5.0-5.5, giving a compound of the formula I, wherein R represents CH(OCH3)2, R1 represents H, R2 represents acetyl, R3 represents OH, R4 represents N(CH3)2, the lines - - - and . . . represent single bonds and the line  represents a double bond,
or optionally,
a compound of the formula I, wherein R represents CH(OCH3)2, R1, R2 and R3 represent H, R4 represents N(CH3)2, the lines - - - and . . . represent double bonds and the line  represents a single bond, is subjected to the oxidation reaction in the manner described under D, and the obtained compound of the formula I, wherein R represents CH(OCH3)2, R1, R2 and R3 represent H, R4 represents Nxe2x80x94O(CH3)2, the line - - - represents a double bond, the line . . . represents  and the line  represents a single bond,
is optionally subjected
to the catalytic hydrogenation reaction in the manner described under C, giving a compound of the formula I, wherein R represents CH(OCH3)2, R1, R2 and R3 represent H, R4 represents N(CH3)2, the line - - - represents a single bond, the line . . . represents  and the line  represents a single bond,
or optionally,
to the reduction with Zn-powder in the manner described under E, giving a compound of the formula I, wherein R represents CH(OCH3)2, R1 and R2 represent H, R3 represents OH, R4 represents N(CH3)2, the lines - - - and . . . represent single bonds and the line  represents a double bond,
or optionally,
a compound of the formula III, wherein R represents H and the line - - - represents a double bond, is subjected to the catalytic hydrogenation reaction in the manner described under C and the obtained compound of the formula III, wherein R represents H and the line - - - - - represents a single bond,
is optionally subjected
F/ to a mesylating reaction in a solution of pyridine under the addition of 3-5 equivalents of methanesulfochloride at 10xc2x0 C. within 3-5 hours and the obtained compound of the formula III, wherein R represents SO2CH3 and the line - - - represents a single bond, is subjected
G/ to a reaction of elimination of mesylate in a mixture of methanol and 25% NH4OH (2:1) at room temperature within 5 hours and, subsequently, to the methanolysis in the manner described under B1, and the obtained compound of the formula II, wherein R represents CH(OCH3)2, R1 represents H, R2 represents N(CH3)2, the lines - - - , . . . and  represent single bonds, is subjected to the reaction of hydrolysis of acetal in the manner described under B2,
or optionally,
a compound of the formula II, wherein R represents CH(OCH3)2, R1 represents H, R2 represents N(CH3)2, the lines - - - and . . . represent double bonds and the line  represents a single bond, is subjected to the oxidation reaction in the manner described under D and the obtained compound of the formula II, wherein R represents CH(OCH3)2, R1 represents H, R2 represents Nxe2x80x94O(CH3)2, the line - - - represents a double bond, the line . . . represents  and the line  represents a single bond,
is optionally subjected to the reduction reaction in the manner described under E, giving a compound of the formula II, wherein R represents CH(OCH3)2, R1 represents OH, R2 represents N(CH3)2, the lines - - - and . . . represent single bonds and the line  represents a double bond.
According to the present invention the isolation of the products is performed by means of conventional extraction processes from alkaline aqueous solutions by the use of halogenated hydrocarbons such as methylene chloride, chloroform or tetrachloro-methane, followed by evaporation to a dry residue.
The course of the reaction is followed by chromatography on a thin layer of silica gel (Merck 60 F254) in solvent systems methylene chloride-methanol-ammonium hydroxide 25% (90:9:1.5, system A), (90:9:0.5, system A1) or methylene chloride-acetone (8:2, system B) (7:3, system C). If appropriate, the separation of the reaction products and the purification of the products for the purpose of spectral analyses are performed on a silica gel column (Merck 60, 230-400 mesh/ASTM, or 60-230 mesh/ASTM in solvent systems A, B or C). The identification of the novel compounds is performed by UV and NMR spectroscopies and by mass analysis.
The novel compounds show antibacterial activity, however, they can also be used as intermediates for the preparation of new derivatives.
The present invention is illustrated but in no way limited by the following Examples.
Preparation of 2xe2x80x2,4xe2x80x2-diacetyl-desmycosin 20-dimethylacetal
Desmycosin 20-dimethylacetal (20 g, 24.4 mmol) was dissolved in methylene chloride (100 ml), acetanhydride (7.2 ml, 76.2 mmol) was added thereto and it was stirred at room temperature for 1 hour. The reaction mixture was poured into 400 ml of water, alkalized to a pH value of 8.5 and subsequently, after the removal of the organic layer, extracted once more with methylene chloride. The combined extracts were washed with a saturated NaHCO3 solution, dried and evaporated to a dry residue.
Obtained: 19.6 g, 89.0%; Rf (A) 0.68; Rf (B) 0.45; MH+ 902.
Preparation of 2xe2x80x2,4xe2x80x2,4xe2x80x3-triacetyl-desmycosin 20-dimethylacetal
2xe2x80x2,4xe2x80x2-diacetyl-desmycosin 20-dimethylacetal (19.6 g, 21.7 mmol) was dissolved in methylene chloride (700 ml) and 4-(dimethylamino)pyridine (0.54 g, 3.7 mmol), triethylamine (27 ml) and acetanhydride (2.7 ml, 28.5 mmol) were added thereto. The reaction mixture was stirred at room temperature for 2 hours, poured into 1000 ml of water and, after the removal of the organic layer, it was extracted once more with methylene chloride. The combined extracts were dried and evaporated to a dry residue.
Obtained: 19.5 g, 95.1%; Rf (A) 0.90; Rf (B) 0.60; MH+ 944.
Preparation of 3-methansuffonyl-2xe2x80x2,4xe2x80x2,4xe2x80x3-triacetyl-desmycosin 20-dimethylacetal
2xe2x80x2,4xe2x80x2,4xe2x80x3-triacetyl-desmycosin 20-dimethylacetal (3 g, 3.18 mmol) was dissolved in pyridine (9.5 ml) and it was cooled to 10xc2x0 C., whereupon methanesulfochloride (1.57 ml, 12.4 mmol) was gradually added thereto. The reaction solution was stirred for 3 hours at 10xc2x0 C., whereupon it was poured into 250 ml of water, alkalized to a pH value of 9.2 and kept under stirring for 30 minutes. A thick white precipitate was separated by filtration and the still moist precipitate was dissolved in chloroform (60 ml) and washed with a saturated NaCl solution (120 ml). The extract was dried and evaporated to a dry residue.
Obtained: 3.05 g, 94.1%; Rf (A) 0.95; Rf (B) 0.70; MH+ 1022.
Preparation of 2,3-anhydro-desmycosin 20-dimethylacetal
3-methanesulfonyl-2xe2x80x2,4xe2x80x2,4xe2x80x3-triacetyl-desmycosin 20-dimethylacetal (3 g, 2.9 mmol) was dissolved in methanol (60 ml), 25% NH4OH (30 ml) was added thereto and it was stirred at room temperature for 3 hours. The reaction mixture was evaporated to ⅓ of its volume under reduced pressure, extracted with chloroform, dried and evaporated to a dry residue. The crude product was dissolved in methanol (160 ml) and heated at reflux temperature for 6 hours, whereupon methanol was evaporated and the product was dissolved in chloroform (150 ml), washed with a saturated NaHCO3 solution and evaporated to a dry residue.
Obtained: 2.22 g, 94.4%; Rf (A) 0.50; MH+ 800.